The encapsulation of materials may provide beneficial properties. For example, drugs that are encapsulated may provide increased stability, longer duration of action and increased efficiency. For convenience drugs are often encapsulated in solid materials which have a size suitable for injection, that is generally below 200 μm in diameter, and then the process is referred to as a microencapsulation.
Microencapsulation processes may yield microcapsules, microspheres or microparticles. Microcapsules consist of a core and a shell that covers the core. The core may be composed of another polymer than the shell or of another material altogether, e.g. of the active substance itself. The active substance is generally located in the core but may also be located in the outer shell. Microspheres are spherical in shape and have a more homogenous matrix. Microparticle is a more general term than microspheres in that it is not restricted to spherical shapes. Sometimes it can be difficult to distinguish between microcapsules, microspheres and microparticles, and the term microparticles will be used herein with reference to all three classes.
Methods of preparing microparticles in the prior art have been described extensively in both the patent and the scientific literature (see e.g. Jalil R, Nixon J R. Biodegradable poly(lactic acid) and poly(lactide-co-glycolide) microcapsules: problems associated with preparative techniques and release properties. J Microencapsul 1990;7:297-325). They may generally be classified in three types, which are exemplified below in connection with the preparation of microspheres of poly(lactide-coglycolide) (PLGA). PLGA is a well accepted polymer for preparing sustained release microspheres and often the first choice for preparing biocompatible microspheres intended for parenteral administration in humans. Said polymer is not soluble in water.
1. Phase separation techniques using coacervating agents, or non solvents, such as mineral oils and vegetable oils. The active substance, e.g. a polypeptide is first dissolved in the aqueous phase of a water-in-oil emulsion. The polypeptide can also be dispersed directly in the polymer phase as a fine powder. Polymer is precipitated either around the aqueous droplets, or on the polypeptide powder, by the addition of a non-solvent for the polymer, such as silicon oil. Then a hardening agent is added to extract the organic solvent from the micro-spheres. The main disadvantage with said process is the large amount of organic solvent needed for extraction and for washing. The previously used hardening agents including freons, hexane, heptane, cyclohexane and other alkane solvents leave substantial amounts of hardening agents residues in the microspheres and/or necessitate extensive procedures for removing the solvent. Often very large amounts of the second organic solvent are needed and they are often undesirable for health, economical and environmental reasons. Examples in the prior art include heptan (EP 0 052 510), aliphatic fluorinated and fluorohalogenated hydrocarbons sold as FREONS (SE 462 780), and other (U.S. Pat. No. 5,000,886). A further drawback when using e.g. an alkane hardening solvent is that it is flammable. Another drawback is the impact thereof on the environment.
2. Spray drying and spray coating
In spray drying the polymer and the drug are mixed together in a solvent for the polymer. The solvent is then evaporated by spraying the solution. This results in polymeric droplets containing the drug. However, sensitive substances such as proteins can be inactivated during the process due to the elevated temperatures used and the exposure to organic solvent/air interfaces. Further disadvantages include generation of high porosity due to rapid removal of the organic solvent. A variation that has been introduced to avoid these shortcomings is the use of low temperature during microsphere formation (U.S. Pat. No. 5,019,400, WO 90/13780 and U.S. Pat. No. 4,166,800). Microcapsules have been prepared using spray coating of drug-containing microparticles with PLGA polymers (U.S. Pat. No. 4,568,559).
3. Solvent evaporation
In solvent evaporation techniques the polymer is dissolved in an organic solvent which contain the dispersed active drug, the solution then being added to an agitated aqueous outer phase which is immiscible with the polymer. The aqueous outer phase usually contains surfactants to stabilise the oil-in-water emulsion and to prevent agglomeration. The emulsifier used is typically polyvinylalcohol. Emulsifiers are included in the aqueous phase to stabilise the oil-in-water emulsion. The organic solvent is then evaporated over a period of several hours or more, thereby solidifying the polymer to form a polymeric matrix. The solvent can also be extracted by adding the above mentioned suspension to a large volume of water (U.S. Pat. No. 5,407,609).
The final formulation to be used for pharmaceutical applications, especially for parenteral administration, should consist of discrete, non-agglomerated microspheres with the desired size distribution and containing no toxic or in any other way undesirable substances. In order to obtain preparations having the characteristics described above it is necessary to use emulsifiers. The emulsifier can serve several purposes: (1) assist in obtaining the correct droplet size distribution of the emulsion; (2) stabilise the oil-in-water emulsion to avoid coalescence of the droplets; and (3) prevent the solidified microspheres from sticking to each other. The most commonly used emulsifier for preparing PLGA microspheres is polyvinyl alcohol. However, since polyvinyl alcohol is listed in the 1976 Register of Toxic Effects of Chemical Substances and is also implicated as carcinogenic when introduced parenterally into animals (“Carcinogenic studies on Water-Soluble and Insoluble Macromolecules”, Archives of Pathology, 67, 589-617, 1959) it is considered undesirable for pharmaceutical preparations administered by injection. This problem has been recognized and attempts of replacing polyvinyl alcohol with other emulsifers can be found in the prior art, for example in U.S. Pat. No. 4,384,975, wherein a carboxylic acid salt surfactant, e.g. sodium oleate was used to stabilise an oil-in-water emulsion. However, despite its drawbacks polyvinyl alcohol is still the most videly used surfactant. However, for the above-mentioned reasons it would be highly desirable to avoid the use of polyvinyl alcohol and other surfactants in microsphere preparations.
Solvent evaporation works well for hydrophobic drugs but for hydrophilic drugs, such as many peptides and proteins, the amount of incorporated drug can be low due to loss of drug to the aqueous phase which is used to extract the organic solvent. Attempts to circumvent this problem include modifying the hydrophilic drug into a less soluble form (WO 96/07399) increasing the viscosity of the inner aqueous solution containing the active drug in a process where a water-in-oil emulsion is first created and the organic solvent then extracted with water (U.S. Pat. No. 4,652,441) and reducing the time available for diffusion (U.S. Pat. No. 5,407,609).
Further, the use of the commonly employed organic solvents, like methylene chloride or ethyl acetate, often results in loss of biological activity for sensitive drugs. Thus, for instance for proteins the three dimensional conformation which is required for biological activity is often lost. Attempts to circumvent this problem includes modification of the active substance into a more stable form (U.S. Pat. No. 5,654,010 and WO 96/40074) keeping the temperature as low as possible during the process (WO 90/13780), and using different protein stabilisers (U.S. Pat. No. 5,589,167, Cleland J L, Jones AJS, “Development of stable protein formulations for microencapsulation in biodegradable polymers”. Proceedings of the International Symposium on Controlled Release of Bioactive Materials 1995;22:514-5). However, proteins are generally sensitive to organic solvents and reducing or eliminating the exposure is highly desirable.
Another disadvantage with the solvent evaporation method is the need for using high shear mixing in order to obtain small microspheres- or nanospheres. This may result in degradation or conformational changes of the active substance, especially if it is a protein which is dependent on a three dimensional conformation for its biological activity. The use of high shear mixing is also energy consuming.
In connection with the prior art it can also be added that processes for preparing microspheres from polymers soluble in water are known from e.g. U.S. Pat. No. 4,822,535 and U.S. Pat. No. 5,578,709. In said processes two mutually immiscible aqueous liquid phases are used, of which one is solidified into microspheres. However, as said, these methods cannot be used for the preparation of microspheres from polymers that cannot be dissolved in water.
The present invention relates to a novel method of encapsulating active substances in biodegradable polymers by which the prior art disadvantages are eliminated or at least essentially reduced. For instance the invention makes it possible to obtain high incorporation efficiency of the active substance in the biodegradable polymer and/or to accomplish smaller microparticles or even nano-particles containing highly active doses of the active substances. Furthermore, the amounts of organic solvents are highly reduced. As compared to previously used methods the invention also enables a reduction of the energy input required to obtain micro- or nanoparticles.